Allylsulfoxide enzyme inhibitors

ABSTRACT

Organic sulfoxides having a latent allyl group bound to the sulfur are enzyme inhibitors of the suicide or K cat  type.

This is a division of application Ser. No. 498,344, filed May 26, 1983,now U.S. Pat. No. 4,491,590, which in turn is a division of applicationSer. No. 349,809, filed Feb. 18, 1982, now U.S. Pat. No. 4,400,395,which in turn is a division of application Ser. No. 147,656, filed May7, 1980, now U.S. Pat. No. 4,332,813, which in turn, is acontinuation-in-part of application Ser. No. 066,603, filed Aug. 15,1979, now abandoned.

BACKGROUND OF THE INVENTION

This invention is concerned with a novel class of enzyme inhibitors ofthe suicide or K_(cat) type in which the latent reactive group is anallylsulfoxide which is in reversible equilibrium with an allylsulfenate: ##STR1##

Suicide enzyme inhibitors are substances bearing a latent reactive groupthat is unmasked by the target enzyme itself, and which after beingunmasked, immediately reacts with the enzyme in an irreversible manner,inactivating it. Enzyme inhibitors of the suicide type are known in theart but until now almost invariably have employed a Michael acceptor asthe reactive species and these are described by Walsh in HorizonsBiochem. Biophys., 3, 36-81 (1977).

The allylsulfoxide-allyl sulfenate equilibrium of reaction scheme (A) isalso known in the art and has been studied as an interesting chemicalreaction by Mislow et al., J. Amer. Chem. Soc., 90, 4869 (1968); 92,2100 (1970) and Evans et al., J. Amer. Chem. Soc., 94, 3672 (1972).Generally, allylsulfoxides are unreactive, but allyl sulfenates arehighly reactive electrophiles, and would be expected to capture almostany nucleophile (Nu) in an enzyme that happens to be near it at themoment it is formed: ##STR2##

Usually the nucleophile is one from the protein portion (prostheticgroup) of the enzyme, such as a sulfhydryl, amino, hydroxy, imidazolylor the like. Once the nucleophile is sulfenylated, the enzyme is alteredfrom its native, active form and can no longer function in its intendedrole as a biochemical catalyst.

The allylsulfoxide-allyl sulfenate rearrangement is facilitated by thenature of the R group attached to the sulfur: the stronger the electronwithdrawing nature of R the better, for example, p-nitrophenyl. Stericacceleration of the rearrangement is also provided by bulkyo-substituent such as o-alkyl and o,o'-dialkyl when R issubstituted-phenyl. Bulky groups such as alkyl and chloro substituted onthe carbon chain adjacent to the sulfur atom also provide stericacceleration.

In the present invention, the latency of the allylsulfoxide group isgenerally secured as a β- or γ-halosulfoxide wherein the halo is also β-to the carboxyl group of an α-amino acid which the target enzymerecognizes as a potential substrate. On attack by the enzyme the aminoacid is decarboxylated, and splits out halide ion to produce theallylsulfoxide.

In the present invention, the allyl sulfoxide type of inhibitoradvantageously also is combined with other types of inhibitor in thesame molecule such as the fluoromethyl dopa decarboxylase, histidinedecarboxylase, or the tryptophane decarboxylase inhibitors. In thesecases, the bifunctionality creates inhibitors with great efficiency,doubling the sites for nucleophilic attack, as shown below: ##STR3##

The mechanism of inhibition, I→II→III is that of Kollonitsch et al.,Nature, 274, 906 (1978).

It is, therefore, an object of this invention to provide a group ofnovel organic sulfoxides wherein one of the substituents on the sulfurcarries such other functional group or groups as to be a latent allylgroup which becomes unmasked upon reaction with a target enzyme andwhich function as enzyme inhibitors of the suicide type.

It is another object of this invention to provide a useful tool ofbiochemical research in the form of selective, very active enzymeinhibitors.

It is a further object of this invention to provide means for inhibitingenzymes, both in vitro and in vivo with the novel organic sulfoxides ofthis invention.

It is a still further object to provide a method of treating diseasestates, the progress of which is dependent on the activity of enzymes,which comprises the administration of an effective amount of an enzymeinhibitor of this invention.

It is also an object of this invention to provide pharmaceuticalformulations comprising one or more of the novel enzyme inhibitors ofthis invention.

DETAILED DESCRIPTION OF THE INVENTION

This invention comprises, as one embodiment, a new class of K_(cat) orsuicide enzyme inhibitors, which are organic sulfoxides of formula:##STR4## or a pharmaceutically acceptable salt thereof, wherein X ishydrogen, fluoro, chloro, bromo, iodo, C₂₋₄ alkanoyloxy,toluenesulfonyloxy, benzenesulfonyloxy, C₁₋₃ alkanesulfonyloxy,p-nitrobenzoyloxy, or the like, or ##STR5## Y is hydrogen or ##STR6##and Z is fluoro, chloro, bromo, iodo, C₂₋₄ alkanoyloxy,toluenesulfonyloxy, benzenesulfonyloxy, C₁₋₃ -alkanesulfonyloxy,p-nitrobenzoyloxy, or the like, or ##STR7## or Y and Z taken togetherform ═CH₂ ; with the proviso that one and only one of X, Y and Z is asulfoxide group, and one and only one of X and Z is fluoro, chloro,bromo, iodo, C₂₋₄ alkanoyloxy, toluenesulfonyloxy, benzenesulfonyloxy,C₁₋₃ alkanesulfonyloxy, p-nitrobenzoyloxy, or the like; wherein

R is

(a) phenyl, either unsubstituted or substituted with such as:

(1) nitro,

(2) cyano,

(3) C₁₋₃ alkylsulfonyl,

(4) C₁₋₃ alkoxycarbonyl,

(5) o-C₁₋₃ alkyl,

(6) o,o'-di(C₁₋₃ alkyl), or

(7) di(trifluoromethyl);

(b) trihalomethyl such as

(1) trifluoromethyl, or

(2) trichloromethyl;

(c) 5-6 membered heteroaryl such as

(1) thiazolyl,

(2) imidazolyl,

(3) pyridinyl,

(4) pyrazinyl,

(5) oxazolyl

(6) pyrimidinyl, or

(7) thienyl; and

R¹ is

(a) imidazol-4-yl,

(b) 3,4-dihydroxyphenyl,

(c) aminoethyl,

(d) 5-hydroxyindol-3-yl,

(e) 4-hydroxyphenyl, or

(f) hydrogen.

Many disease states of mammals, including humans, are known to dependfor their progress on the activity or hyperactivity of particularenzymes and treatment of many of these diseases have been devised aroundinhibitors of these enzymes. Accordingly, the novel enzyme inhibitors ofthis invention have utility in the study of certain disease states andin their treatment.

Generally the novel enzyme inhibitors of this invention produce thedesired effect when administered at from 0.1 to about 500 mg/kg bodyweight, preferably at from 1 to about 50 mg/kg of body weight. Thepreferred form of delivery of the instant compounds to domestic animalsis by solution in drinking water or by inclusion in preformulatedfeedstuffs. For human and animal administration, any of the usualpharmaceutical oral forms may be employed such as tablets elixirs,aqueous suspensions or the like comprising from about 0.1 to about 500mg of the compounds of this invention. Sterile solutions(representatively given for human treatment) for injection comprisingfrom about 0.1 to about 500 mg of the compounds of this invention giventwo or four times daily are also suitable means of delivery.

Representative specific members of the new class of suicide enzymeinhibitors are shown in Table I along with the enzyme to be inhibitedand the pharmacological or medical effect to be elicited. In each case,R represents o- or p-nitrophenyl, o- or p-cyanophenyl, o orp-methoxycarbonylphenyl, o- or p-methylsulfonylphenyl,o,p-di(trifluoromethyl)phenyl, trifluoromethyl, trichloromethyl,2-pyrimidinyl, 2-pyridyl, 2-imidazolyl, 2-thienyl, 2-thiazolyl,2-oxazolyl, o-methylphenyl, o-ethylphenyl, o-propylphenyl,o,o-di(methyl)phenyl, o,o-di(ethyl)phenyl, or o,o-di(propyl)phenyl.

                                      TABLE I                                     __________________________________________________________________________                                        USE,                                                                          PHARMACOLOGICAL OR                        INHIBITOR              ENZYME INHIBITED                                                                           MEDICAL EFFECT                            __________________________________________________________________________     ##STR8##              histidine decarboxylase                                                                    antihistamine                              ##STR9##                                                                      ##STR10##             histidine decarboxylase                                                                    antihistamine                              ##STR11##                                                                     ##STR12##             dopa decarboxylase                                                                         antihypertensive antiparkinson when                                           administered with L-dopa                   ##STR13##                                                                     ##STR14##             dopa decarboxylase                                                                         antihypertensive antiparkinson when                                           administered with L-dopa                   ##STR15##                                                                     ##STR16##             ornithine decarboxylase                                                                    anti-psoriasis anti-arthritis                                                 anti-cancer                                ##STR17##                                                                     ##STR18##             ornithine decarboxylase                                                                    anti-psoriasis anti-arthritis                                                 anti-cancer                                ##STR19##                                                                     ##STR20##                                                                     ##STR21##             tryptophan decarboxylase                                                                   antiserotonin                              ##STR22##                                                                     ##STR23##             tryptophan decarboxylase                                                                   antiserotonin                              ##STR24##                                                                     ##STR25##             tyrosine hydroxylase                                                                       antihypertensive                           ##STR26##                                                                     ##STR27##             tyrosine hydroxylase                                                                       antihypertensive                           ##STR28##                                                                    __________________________________________________________________________

The novel process for preparing the novel compounds of this inventioncomprises oxidation of an aromatic thio compound of structure: ##STR29##wherein X¹ is hydrogen, fluoro, chloro, bromo, iodo, C₂₋₄ alkanoyloxy,toluenesulfonyloxy, benzenesulfonyloxy, C₁₋₃ alkanesulfonyloxy,p-nitrobenzoyloxy, or the like, or ##STR30## Y¹ is hydrogen or ##STR31##and Z¹ is fluoro, chloro, bromo, iodo, C₂₋₄ alkanoyloxy,toluenesulfonyloxy, benzenesulfonyloxy, C₁₋₃ alkanesulfonyloxy,p-nitrogenzoyloxy, or the like, or ##STR32## or Y and Z taken togetherform ═CH₂ ; with the proviso that one and only one of X, Y and Z is athio group, and one and only one of X and Z is fluoro, chloro, bromo,iodo, C₂₋₄ alkanoyloxy, toluenesulfonyloxy, benzenesulfonyloxy, C₁₋₃alkanesulfonyloxy, p-nitrobenzoyloxy, or the like; and R, and R¹ are aspreviously defined with the exception that any of the substituents whichare sensitive to the conditions of oxidation of sulfide to sulfoxidecarry protective groups.

The oxidizing agent is such as 1-chlorobenzotriazole, H₂ O₂ /V₂ O₅, SO₂Cl₂ /H₂ O/silica gel, Cl₂, Br₂, NaIO₄, acetyl nitrate, Tl(NO₃)₃, or aperacid such as m-chlorperbenzoic acid, preferably the latter. Theoxidation with a peracid is conducted at temperatures from -70° C. toabout 30° C., preferably at about 0°-25° C., in an organic solvent suchas an aromatic solvent, for example benzene, toluene or the like; or achlorinated hydrocarbon such as tetrachloroethylene, chloroform,methylene chloride or the like, for times of a few minutes to about 4hours.

After the oxidation is substantially complete, any protective groupspresent are removed by standard procedures such as treatment with astrong organic acid such as trifluoroacetic acid to removet-butyloxycarbonyl groups from amines and to cause deesterification;strong mineral acids to remove trityl groups from amines; and strongbases such as sodium hydroxide or potassium hydroxide to saponifyesters.

EXAMPLE 1 2-Fluoromethyl-3-(p-nitrophenylsulfinyl)histidinehydrochloride

Step A: Preparation of N.sub.α -phthaloyl-2-fluoromethylhistidine methylester (1·I) ##STR33##

2-Fluoromethyl histidine methyl ester, 2.01 g (10 mmoles) and 1.48 g ofphthalic anhydride (10 mmoles) are ground together in a mortar and thenheated together 2 hours at 150° C. to form compound 1·I.

Step B: Preparation of N_(Im) -trityl-N.sub.α-phthaloyl-2-fluoromethylhistidine methyl ester (1·II) ##STR34##

Compound 1·I, 331 mg (1 mmole), is treated with 278.5 mg (1 mmole) oftrityl chloride in 25 ml of DMF containing 139 μl of triethylamine (1mmole) overnight. The DMF is pumped off in vacuo and the residue istaken up in benzene, is washed with dilute aqueous sodium bicarbonatethree times, then brine, then dried over K₂ CO₃. Filtration andevaporation of the solvent affords 1·II.

Step C: Preparation of N_(Im) -trityl-N.sub.α-phthaloyl-2-fluoromethyl-2'-trimethylsilyl histidine methyl ester(1·III) ##STR35##

Lithium 2,2,6,6-tetramethyl piperidide (LiTMP) (1 mmole) is prepared asfollows: To 141 mg of TMP (1 mmole) in 10 ml of THF at -18° C. under N₂is added 1 mmole of n-butyllithium. The solution is then aged 15 minutesat 0° C. To it is then added a solution of compound 1·II, 573 mg (1mmole), in 10 ml of THF at -78° under N₂. The reaction mixture is aged30 minutes at -78° C. and 10 minutes at 0° C. and then treated at -78°C. with a solution of 127 μl (1 mmole) of trimethyl silyl chloride in 2ml of THF. The reaction is allowed to warm to room temperature over 30minutes. The solution of 1·III thus obtained is used directly in thenext step.

Step D: Preparation of N_(Im) -trityl-N.sub.α-phthaloyl-2-fluoromethyl-3-(p-nitrophenylthio)-2'-trimethylsilylhistidinemethyl ester (1·IV) ##STR36##

The solution of 1·III from Step C is treated with a second mmole ofLiTMP as before. Then at -78° C. this is added to a suspension of 308 mg(1 mmole) of very finely ground bis-(p-nitrophenyl)disulfide in 25 ml ofTHF. With vigorous stirring, this reaction mixture is allowed to warm toroom temperature over 20 minutes, and then refluxed for 30 minutes. Thesolvent is evaporated, and the residue is taken up in 30 ml of benzene,washed twice with 1N aqueous NaOH, then with dilute aqueous HCl, thenwith brine. After drying over MgSO₄, filtration and evaporation of thesolvent, there is obtained compound 1·IV.

Step E: Preparation of N_(Im) -trityl-N.sub.α-phthaloyl-2-fluoromethyl-3-(p-nitrophenylsulfinyl)-2'-trimethylsilylhistidinemethyl ester (1·V) ##STR37##

To 798 mg of compound 1·IV (1 mmole) in 25 ml of CH₂ Cl₂ is addeddropwise over 1 hour a solution of 203 mg of 85% pure MCPBA (net 172.6mg; 1 mmole) in 20 ml of CH₂ Cl₂. The solution is aged 30 minutes at 25°C. and washed successively with aqueous NaHCO₃ and brine. Evaporationgives compound 1·V.

Step F: Preparation of N_(Im) -trityl-N.sub.α-phthaloyl-2-fluoromethyl-3-(p-nitrophenylsulfinyl)-histidine (1·VI)##STR38##

Compound 1·V, 814 mg (1 mmole), is heated 2 hours at 50° C. with 30 mlof 2N NaOH in 3:2 (v/v) H₂ O--MeOH with stirring. Water, 50 ml, is thenadded, the pH is adjusted to 2.0 with aqueous HCl, and the product isextracted with 3×30 ml of CH₂ Cl₂. The organic extracts are combined,washed with brine, dried with MgSO₄, filtered and evaporated to providecompound 1·VI.

Step G: Preparation of N_(Im)-trityl-2-fluoromethyl-3-(p-nitrophenylsulfinyl)histidine (1·VII)##STR39##

The phthaloyl group is removed by refluxing 728 mg of compound 1·VI (1mmole) with 96 mg of hydrazine hydrate (3 mmoles) in 20 ml of EtOH for 2hours. Aqueous NaOH, 1 mmole, is added, and the reaction mixture ispumped to dryness at 0.1 Torr. Toluene is added and pumped off at 0.1Torr to remove traces of hydrazine. The product is separated fromphthaloyl hydrazide by taking it up in aqueous NaHCO₃ and washing withCH₂ Cl₂. Evaporation of the water affords compound 1·VII as the sodiumsalt.

Step H: Preparation of 2-fluoromethyl-3-(p-nitrophenylsulfinyl)histidinehydrochloride (1·VIII) ##STR40##

Compound 1·VIII, 598 mg (620 mg as the Na salt) of (1 mmole), isrefluxed 3 hours with 25 ml of 6N HCl in MeOH containing 5% by volume ofwater to remove the trityl group. The reaction mixture is evaporated todryness in vacuo, triturated with ether, taken up in ethanol, filteredand evaporated to afford compound 1·VIII.

EXAMPLE 2 2-([1-Fluoro-2-p-nitrophenylsulfinyl]ethyl)histidinehydrochloride

Step A: Preparation of N-benzylidene histidine methyl ester (2·I)##STR41##

Histidine methyl ester, 185 mg (1 mmole), is treated with 1.06 g (1mmole) of benzaldehyde in 25 ml of CHCl₃ for 3 hours. MgSO₄, 0.2 g, isadded for the last hour. The mixture is filtered and the solvent isevaporated, to leaving compound 2·I as a residue.

Step B: Preparation of N_(Im) -trityl-N.sub.α -benzylidene histidinemethyl ester (2·II) ##STR42##

Compound 2·I, 273 mg (1 mmole) is treated with 278.5 mg (1 mmole) oftrityl chloride in 25 ml of DMF containing 139 μl of triethylamine (1mmole) overnight. The DMF is pumped off in vacuo and the residue, takenup in benzene, is washed with dilute aqueous sodium bicarbonate threetimes, then brine, then dried with K₂ CO₃. Filtration and evaporation ofthe solvent afford compound 2·II.

Step C: Preparation of N_(Im) -trityl-N.sub.α-benzylidene-2-(1-hydroxyethyl)histidine methyl ester (2·III) ##STR43##

Compound 2·II, 516 mg (1 mmole), in 20 ml THF under N₂ at -78° C., istreated with 1 mmole of phenyllithium for 1 minute, forming the anion.Acetaldehyde, 44 mg (1 mmole) in 1 ml of THF is then added, and themixture is allowed to warm slowly to room temperature. After evaporationof the solvent and replacement with CHCl₃, the sample is washed withwater, then brine, then dried with K₂ CO₃ and filtered. The filtrate isused directly in the next step.

Step D: Preparation of N_(Im) -trityl-N.sub.α-benzylidene-2-(1-tosyloxyethyl)histidine methyl ester (2·IV) ##STR44##

To the solution of compound 2·III from Step C is added 0.5 ml ofpyridine and then 190.5 mg (1 mmole) of tosyl chloride. After 30 minutesat 25° C. the solution is washed three times with water, once withbrine, and dried with K₂ CO₃ and filtered. The filtrate is used directlyin the next step.

Step E: Preparation of N_(Im) -trityl-N.sub.α -benzylidene-2-vinylhistidine methyl ester (2·V) ##STR45##

The solution of compound 2·IV from Step D is evaporated, taken up inbenzene and refluxed 3 hours with 124 mg (1 mmole) of diazabicyclononane(DBN). The solution is washed twice with water, once with brine, anddried with K₂ CO₃. Filtration and evaporation afford compound 2·V.

Step F: Preparation of N_(Im) -trityl-2-vinylhistidinemethyl ester(2·VI) ##STR46##

Compound 2·V from Step E is taken up in anhydrous ether and addeddropwise over 10 minutes to a solution of 190 mg of p-TSA·H₂ O (1 mmole)in 20 ml of ether. The toslyate salt of 2·VI precipitates out. It iscollected by decantation, and stirred with ether and aqueous sodiumbicarbonate. The ether phase is separated from the aqueous layer, washedwith brine, dried with K₂ CO₃, filtered and evaporated to yield compound2·VI.

Step G: Preparation of N_(IM) -trityl-2-vinyl-N.sub.α -BOC-histidinemethyl ester (2·VII) ##STR47##

Compound 2·VI, 454 mg (1 mmole), is stirred with 218 mg of (BOC)₂ O in30 ml of CH₂ Cl₂ for 3 hours at 25° C. and then washed successively withwater and brine, and dried with K₂ CO₃. Filtration and evaporationprovides compound 2·VII.

Step H: Preparation of N_(Im)-trityl-2-([1-chloro-2-p-nitrophenylthio]ethyl)-N.sub.α -BOC-histidinemethyl ester (2·VIII) ##STR48##

To 554 mg (1 mmole) of compound 2·VIII in 25 ml of CH₂ Cl₂ at -18° C. isadded dropwise over 30 minutes a solution of 190 mg (1 mmole) ofp-nitrophenylsulfenyl chloride in 10 ml of CH₂ Cl₂. The reaction is aged30 minutes at room temperature and evaporated to afford compound 2·VIII.

Step I: Preparation of N_(Im)-trityl-2-([1-fluoro-2-p-nitrophenylthio]ethyl)-N.sub.α -BOC-histidinemethyl ester (2·IX) ##STR49##

Compound 2·VIII, 744 mg (1 mmole), is stirred overnight in 30 ml of dryacetonitrile with 127 mg (1 mmole) of silver fluoride. The precipitatedsilver chloride is separated by centrifugation and the solvent isevaporated to afford compound 2·IX.

Step J: Preparation of N_(Im)-trityl-2-([1-fluoro-2-p-nitrophenylsulfinyl]ethyl)-N.sub.α-BOC-histidine methyl ester (2·X) ##STR50##

To 727 mg of compound 2·IX (1 mmole) in 25 ml of CH₂ Cl₂ at 0° C. isadded dropwise over 1 hour a solution of 172.6 mg (1 mmole); actually203 mg of 85% pure) of MCPBA in 20 ml of CH₂ Cl₂. The solution is aged30 minutes at 25° C. and washed successively with aqueous NaHCO₃ andbrine. Evaporation gives compound 2·X.

Step K: Preparation of2-([1-fluoro-2-p-nitrophenylsulfinyl]ethyl)histidine hydrochloride(2·XI) ##STR51##

Compound 2·X, 745 mg, is treated with 60 mg of KOH (1.05 mmole) in 5 mlof MeOH for 2 hours at 45° C. to saponify the methyl ester. Then 25 ml6N HCl in MeOH containing 5% by volume of water is added and the mixtureis refluxed 3 hours to remove the trityl group. The reaction mixture isevaporated to dryness in vacuo, triturated with ether to remove tritylalcohol, and then taken up in 10 ml of ethanol. The KCl is filtered andthe filtrate evaporated to afford compound 2·XI.

EXAMPLE 3 2-(p-nitrophenylsulfinyl)-3-chlorohistidine hydrochloride

Step A: Preparation of N_(Im) -trityl imidazole-4-carboxaldehyde (3·I)##STR52##

Imidazole 4-carboxaldehyde, 96 mg (1 mmole), is treated with 278.5 mg (1mmole) of trityl chloride in 25 ml of DMF containing 139 μl oftriethylamine (1 mmole) overnight. The DMF is pumped off in vacuo andthe residue, taken up in benzene, is washed with dilute aqueous NaHCO₃three times, then brine, then dried with K₂ CO₃, filtered and evaporatedto afford compound 3·I.

Step B: Preparation of N-BOC-S-p-nitrophenyl cysteine methyl ester(3·II) ##STR53##

Cysteine N-BOC methyl ester, 235 mg (1 mmole), is treated with 24 mg ofNaH (1 mmole) in dioxane with 20 mg of 15-crown-5, and then with 158 mgof p-chloronitrobenzene (1 mmole) at 100° C. for five hours. The dioxaneis evaporated in vacuo, leaving crude 3·II.

Step C: Preparation of S-p-nitrophenyl cysteine methyl ester (3·III)##STR54##

Crude II from Step B is taken up in 2 ml anisole and then treated at 0°C. for 11 minutes with 10 ml of TFA. The TFA and anisole are pumped offat 0.1 Torr at 30° C. and the residue partitioned between chloroform andaqueous 1N HCl. The aqueous layer is made to pH 9.0 with NaOH andextracted with CH₂ Cl₂, providing a solution of compound 3·III.

Step D: Preparation of N-benzylidene S-p-nitrophenyl cysteine methylester (3·IV) ##STR55##

Compound 3·III, 256 mg (1 mmole), in 10 ml of CH₂ Cl₂, is treated with106 mg of benzaldehyde (1 mmole) for two hours at 25° C., then for athird hour with added MgSO₄ (100 mg). The solution is filtered andevaporated, affording compound 3·IV.

Step E: Preparation of N_(Im) -trityl N.sub.α -benzylidene2-(p-nitrophenylthio)-3-hydroxy histidine methyl ester (3·V) ##STR56##

Compound 3·IV, 344 mg (1 mmole), is treated at -78° in 10 ml of THF withone equivalent of PhLi. After one minute, compound 3·I, 338 mg (1mmole), is added, and the reaction allowed to warm to room temperatureover 30 minutes. The solvent is evaporated and the residue is taken upin ether, washed with water and brine, dried with K₂ CO₃, filtered andevaporated to afford compound 3·V.

Step F: Preparation of N_(Im) -trityl N.sub.α -BOC2-(p-nitrophenylthio)-3-hydroxy histidine methyl ester (3·VI) ##STR57##

Compound 3·V, 682 mg (1 mmole), in 5 ml of anhydrous ether is addeddropwise over 10 minutes to a solution of 190 mg of p-TSA·H₂ O (1 mmole)in 20 ml of ether. The tosylate salt of the free N.sub.α amine iscollected by decantation, stirred with CH₂ Cl₂ and aqueous NaHCO₃. Theorganic layer is dried over K₂ CO₃. The solution is filtered and treatedwith 218 mg of BOC₂ O for three hours at 25° C. and then washedsuccessively with water and brine, and dried with K₂ CO₃. Afterfiltration and evaporation, compound 3·VI is obtained.

Step G: Preparation of N_(Im) -trityl N₆₀ -BOC2-(p-nitrophenylthio)-3-chloro histidine methyl ester (3·VII) ##STR58##

Compound 3·VI, 694 mg (1 mmole), is treated with 5 ml of pure SOCl₂ forone hour and then pumped in vacuo, affording compound 3·VII.

Step H: Preparation of N_(Im) -trityl N.sub.α -BOC2-(p-nitrophenylsulfinyl)-3-chloro histidine methyl ester (3·VIII)##STR59##

To 713 mg of compound 3·VII (1 mmole) in 25 ml of CH₂ Cl₂ at 0° C. isadded dropwise over one hour a solution of 172.6 mg (1 mmole; actually203 mg of 85% pure) of MCPBA in 20 ml of CH₂ Cl₂. The solution is aged30 minutes at 25° C. and washed successively with aqueous NaHCO₃ andbrine to afford, after evaporation, compound 3·VIII.

Step I: Preparation of N_(Im) -trityl N.sub.α -BOC2-(p-nitrophenylsulfinyl)-3-chloro histidine (3·IX) ##STR60##

Compound 3·VIII, 729 mg (1 mmole), is treated with 60 mg of KOH in 5 mlof MeOH for two hours at 45° C. to saponify the methyl ester, affordingcompound 3·IX in solution.

Step J: Preparation of 2-(p-nitrophenylsulfinyl)-3-chloro histidinehydrochloride (3·X) ##STR61##

To the solution of compound 3·IX from Step I is added 25 ml of 6N HCl inmethanol+5% water. The mixture is refluxed three hours to remove thetrityl and BOC groups. The product 3·X is isolated by evaporating themethanol, triturating with ether, taking it up in 10 ml of ethanol,filtration and evaporation.

The pharmaceutical formulations of this invention are illustrated in thefollowing Examples which are meant to be illustrative only and notlimiting with respect to type of formulation, nature of pharmaceuticalcarrier or proportions of ingredients.

EXAMPLE 4

    ______________________________________                                                        TABLET                                                                        Per tablet, mg.                                               ______________________________________                                        Levodopa          250                                                         2-Fluoromethyl-3-(4-nitro-                                                                      25                                                          phenylsulfinyl)-3-(3,4-                                                       dihydroxyphenyl)alanine                                                       Lactose           79.0                                                        Starch, corn      65.0                                                        Hydroxypropyl cellulose                                                                         8.0                                                         (as 2% in ethanol)                                                            Add:                                                                          Starch, corn      55.0                                                        Guar gum          55.0                                                        Magnesium stearate                                                                              4.0                                                         ______________________________________                                    

The first four components are reduced to a fine powder by milling andremixing. The mixture is granulated with the hydroxypropyl cellulosesolution. The wetted mass is passed through a No. 10 stainless steelscreen and dried in the dark at 100° F. The dried granules are passedthrough a No. 20 stainless steel screen, and the additional quantity ofcorn starch, guar gum and magnesium stearate added. The mixture iscompressed using a 1/2" standard curvature punch into tablets and thetablet may be coated with a conventional protective film containingvarious types of cellulose polymers, dyes and opacifying agents.

EXAMPLE 10 Injectable Preparation

2-(p-Nitrophenylsulfinyl)-3-chlorohistidine hydrochloride 25 mg.

Pyrogen fee water to 1 ml.

Sterilize by filtration and seal under nitrogen.

What is claimed is:
 1. A compound of the structural formula: ##STR62##or a pharmaceutically acceptable salt thereof wherein X is hydrogen,fluoro, chloro, bromo, iodo, C₂₋₄ alkanoyloxy, toluenesulfonyloxy,benzenesulfonyloxy, C₁₋₃ alkanesulfonyloxy, p-nitrobenzoyloxy, or##STR63## Y is hydrogen or ##STR64## and Z is fluoro, chloro, bromo,iodo, C₂₋₄ alkanoyloxy, toluenesulfonyloxy, benzenesulfonyloxy, C₁₋₃alkanesulfonyloxy, p-nitrobenzoyloxy, or ##STR65## or Y and Z takentogether form ═CH₂ ; with the proviso that one and only one of X, Y andZ is a sulfoxide, and one and only one of X and Z is halo;wherein Ris(1) phenyl, (2) nitrophenyl, (3) cyanophenyl, (4) C₁₋₃alkylsulfonylphenyl, (5) C₁₋₃ alkoxycarbonylphenyl, (6)di(trifluoromethyl)phenyl, (7) o-C₁₋₃ alkylphenyl, or (8) o,o-di(C₁₋₃alkyl)phenyl, and R¹ isaminoethyl, or hydrogen.
 2. The compound of claim1 which is2,5-diamino-2-fluoromethyl-3-(p-nitrophenylsulfinyl)pentanoicacid; or2,5-diamino-2-([1-fluoro-2-p-nitrophenylsulfinyl]ethyl)pentanoic acid;ora pharmaceutically acceptable salt thereof.
 3. A pharmaceutical enzymeinhibiting composition comprising a pharmaceutical carrier and aneffective enzyme inhibiting amount of a compound of structural formula:##STR66## or a pharmaceutically acceptable salt thereof wherein X ishydrogen, fluoro, chloro, bromo, iodo, C₂₋₄ alkanoyloxy,toluenesulfonyloxy, benzenesulfonyloxy, C₁₋₃ alkanesulfonyloxy,p-nitrobenzoyloxy, or ##STR67## Y is hydrogen or ##STR68## and Z isfluoro, chloro, bromo, iodo, C₂₋₄ alkanoyloxy, toluenesulfonyloxy,benzenesulfonyloxy, C₁₋₃ alkanesulfonyloxy, p-nitrobenzoyloxy, or##STR69## or Y and Z taken together form ═CH₂ ; with the proviso thatone and only one of X, Y and Z is a sulfoxide, and one and only one of Xand Z is halo;wherein R is(1) phenyl, (2) nitrophenyl, (3) cyanophenyl,(4) C₁₋₃ alkylsulfonylphenyl, (5) C₁₋₃ alkoxycarbonylphenyl, (6)di(trifluoromethyl)phenyl, (7) o-C₁₋₃ alkylphenyl, or (8) o,o-di(C₁₋₃alkyl)phenyl, and R¹ isaminoethyl, or hydrogen.
 4. The pharmaceuticalcomposition of claim 3 wherein the compoundis2,5-diamino-2-fluoromethyl-3-(p-nitrophenylsulfinyl)pentanoic acid; or2,5-diamino-2-([1-fluoro-2-p-nitrophenylsulfinyl]ethyl)pentanoic acid;ora pharmaceutically acceptable salt thereof.
 5. A method of inhibitingenzymes in a patient in need of such treatment which comprises theadministration of an enzyme inhibitory amount of a compound ofstructural formula: ##STR70## or a pharmaceutically acceptable saltthereof wherein X is hydrogen, fluoro, chloro, bromo, iodo, C₂₋₄alkanoyloxy, toluenesulfonyloxy, benzenesulfonyloxy, C₁₋₃alkanesulfonyloxy, p-nitrobenzoyloxy, or ##STR71## is hydrogen or##STR72## and Z is fluoro, chloro, bromo, iodo, C₂₋₄ alkanoyloxy,toluenesulfonyloxy, benzenesulfonyloxy, C₁₋₃ alkanesulfonyloxy,p-nitrobenzoyloxy, or ##STR73## or Y and Z taken together form ═CH₂ ;with the proviso that one and only one of X, Y and Z is a sulfoxide, andone and only one of X and Z is halo;wherein R is(1) phenyl, (2)nitrophenyl, (3) cyanophenyl, (4) C₁₋₃ alkylsulfonylphenyl, (5) C₁₋₃alkoxycarbonylphenyl, (6) di(trifluoromethyl)phenyl, (7) o-C₁₋₃alkylphenyl, or (8) o,o-di(C₁₋₃ alkyl)phenyl, and R¹ isaminoethyl, orhydrogen.
 6. The method of claim 5 wherein the compoundis2,5-diamino-2-fluoromethyl-3-(p-nitrophenylsulfinyl)pentanoic acid; or2,5-diamino-2-([1-fluoro-2-p-nitrophenylsulfinyl]ethyl)pentanoic acid;ora pharmaceutically acceptable salt thereof.